The incidence and geographic distribution of Lyme disease in the U.S. has increased steadily since its first description in 1977. Efforts to stem the spread of the disease through controlling the population of its tick vector and/or the mouse reservoirs of the disease have met with only limited success. The only approved human vaccine to protect against Lyme disease was recently removed from the market by its manufacturer further highlighting the need for new approaches to controlling the disease. In this project, we propose the development of an orally-available vaccine targeted towards the mouse and tick reservoirs of the disease. This proposal combines the expertise of Foodsource Lure Corp, a manufacturer of animal baits that is currently involved in the development of wildlife-targeted vaccines for rabies and the laboratory of Dr. Linden Hu at Tufts University. Dr. Hu's laboratory has developed an oral vaccine for mice using a vaccinia virus vector expressing the B. burgdorferi protein outer surface protein A (OspA). Vaccination of mice protects them against infection with B. burgdorferi by feeding ticks as well as protects uninfected ticks from acquiring infection from vaccinated but infected mice giving the vaccine two potential mechanisms for decreasing environmental persistence of B. burgdorferi. In phase I of this project, we developed a stable, highly palatable system for delivering the vaccine to mice and shown that it is immunogenic and protective. We also developed and field tested a strategy for bait delivery that maximizes penetration of the vaccine into rodent populations. In this phase II STTR proposal, we will focus on translating our phase I research towards the development of a commercial product. The next step towards commercialization is receiving USDA regulatory approval to produce and distribute the vaccine in field trials. To this end, we propose three aims designed to produce the information required for a USDA application for an animal vaccine. Our goal is to: 1) create the master seed virus (MSV) that will be used for all subsequent development; 2. test the MSV for stability and purity in accordance with stringent USDA standards for commercial products; 3. produce safety data for the vaccine in target and non-target animals; 4. perform simulated field trials in containment that will generate data confirming environmental safety and feasibility of the animal vaccine strategy. At the completion of this phase II proposal, we will have completed all requirements to submit our application for regulatory approval to USDA. Development of reservoir targeted vaccine strategies for controlling vector transmitted diseases will be an important addition to the armamentarium for prevention of human Lyme disease.